H3.3 incorporation in muscle-specific gene loci safeguards myogenic cell identity

Description

Abstract Chromatin architecture impacts on gene expression and defines cell lineage specification. Although epigenetic processes in myogenesis start to be unveiled, the role of the histone variant H3.3, which is implicated in epigenetic memory, is poorly understood. By combining studies using a HIRA-KO myoblast cell line and the Pax7CreErt2;Hirafl/fl mouse model we showed that myoblasts and skeletal muscle stem cells lacking HIRA lose Pax7, Myf5 and Myod1 expression, consistent with the loss of PAX7-positive cells and impaired muscle regeneration. Mechanistically, in the absence of HIRA, H3.3 enrichment in regulatory regions of myogenic genes is decreased and correlates with the loss of the transcriptional permissive histone mark H3K27ac. Moreover, loss of myogenic identity in HIRA mutant cells is associated with the atypical expression of alternative lineage genes which is linked with the increased enrichment of H3K4me3 at promoters. Consistently, HIRA is required for myogenic development, with decreased muscle progenitor cells and muscle size in Pax3Cre;Hirafl/fl embryos. In order to investigate gene expression and motif accessibility at the single cell level, we performed multiome analysis (scRNA-seq coupled with scATAC-seq) and observed distinct clusters of muscle stem cells that respond differently in the context of HIRA loss-of-function. Taken together, our data demonstrate that nucleosome incorporation of H3.3 by HIRA safeguards myogenic gene expression and represses that of alternative lineage genes.